Shiga toxin-producing strains of enterohaemorrhagic Escherichia coli (EHEC) are a class of pathogenic E. coli responsible for numerous food- and water-borne disease outbreaks although other transmission routes are also evident. EHEC causes a range of illnesses from non-bloody diarrhoea through haemorrhagic colitis to severe life-threatening haemolytic uremic syndrome (reviewed in (Nataro and Kaper, Diarrheagenic Escherichia coli, Clin Microbiol Rev 11: 142-201 (1998); Paton and Paton, Pathogenesis and diagnosis of Shiga toxin-producing Escherichia coli infections, Clin Microbiol Rev 11: 450-79 (1998)). Strains of EHEC O157:H7, the most common serotype causing human disease, remain an important cause of zoonotic infection throughout Northern Europe, North America and Japan in particular.
Attachment to the intestinal epithelium surfaces is an important initial step in pathogenesis of EHEC. The EHEC intimate adherence and A/E lesion formation in vitro and in vivo is mediated by the locus of enterocyte effacement (LEE) pathogenicity island, which encodes a type III protein secretion system. One of the LEE-encoded type III secreted proteins (Tir) is translocated into the host cell where it forms a hairpin structure in the host cell plasma membrane with an extracellular loop and two amino- and carboxy-terminal transmembrane domains (Kenny et al., Enteropathogenic E. coli (EPEC) transfers its receptor for intimate adherence into mammalian cells, Cell 91: 511-20 (1997); de Grado et al., Identification of the intimin-binding domain of Tir of enteropathogenic Escherichia coli, Cell Microbiol 1: 7-17 (1999)). The extracellular loop of Tir interacts directly with the LEE-encoded outer membrane protein intimin, thus anchoring the bacteria tightly to the host cell (de Grado et al., (1999)). The cytoplasmic domains of Tir binds to the host cytoskeletal and signalling proteins and initiate actin polymerization at the site of bacterial attachment (Goosney, et al., Recruitment of cytoskeletal and signaling proteins to enteropathogenic and enterohemorrhagic Escherichia coli pedestals, Infect Immun 69: 3315-22 (2001); Gruenheid et al., Enteropathogenic E. coli Tir binds Nck to initiate actin pedestal formation in host cells, Nat Cell Biol 3: 856-9 (2001) Campellone K G et al., EspFU is a translocated EHEC effector that interacts with Tir and N-WASP and promotes Nck-independent actin assembly, Dev Cell. 2004 August; 7(2):217-28). This results in the formation of actin pedestal structures underneath adherent bacteria.
Although a substantial amount of data has been generated in recent years regarding the interaction of E. coli O157:H7 with host cells, so far type III secretory proteins are the only O157:H7 virulence determinants demonstrated to play a direct role in attachment to intestinal mucosa in vivo. However, their role in intimate adherence is likely to be limited to later stages of infection (Donnenberg et al., The role of the eae gene of enterohemorrhagic Escherichia coli in intimate attachment in vitro and in a porcine model, J Clin Invest 92: 1418-24 (1993); McKee et al., The role of the eae gene of enterohemorrhagic Escherichia coli in intimate attachment in vitro and in a porcine model, J Clin Invest 92: 1418-24 (1995); Tzipori et al., The role of the eaeA gene in diarrhea and neurological complications in a gnotobiotic piglet model of enterohemorrhagic Escherichia coli infection, Infect Immun 63: 3621-7 (1995)) and factors conferring initial interaction of EHEC with intestinal epithelium remain to be clearly defined. Some people believe that EspA-containing surface appendages are important in initiating contact between EHEC and their target cells. After initial contact there is a gradual reduction in these filaments which are later replaced by tighter attachment mediated by intimin (Ebel et al., Initial binding of Shiga toxin-producing Escherichia coli to host cells and subsequent induction of actin rearrangements depend on filamentous EspA-containing surface appendages, Mol Microbiol 30: 147-61 (1998)). Although the factors responsible may not be fully defined, it is clear that the molecular interactions of E. coli O157:H7 with intestinal epithelium are complex and multiphasic and likely involve multiple types of ligand-receptor contacts during the course of colonization.
Recently, evidence has been presented to support a role for E. coli flagella in adherence to epithelium, not merely via motility/chemotaxis, but directly as an adhesin (Giron et al., The flagella of enteropathogenic Escherichia coli mediate adherence to epithelial cells, Mol. Microbiol. 44: 361-379 (2002)). Specifically, purified H6 and H2 flagella of EPEC bound human epithelial cells, as assessed non-quantitatively by immunofluorescence, and anti-H6 flagella antibodies inhibit adherence of EPEC strain E. coli O127:H6 (E2348/69). The H6 fliC mutant show a 60% reduction in adherence and introduction of fliC gene from the EPEC strain into a K-12 strain conferred adherence reminiscent of localized adherence. Hence flagella are implicated in pathogenicity of EPEC for which roles in initial adherence and microcolony formation is proposed.
A recent article demonstrates that flagella deficient Shiga-toxigenic E. coli O113:H21 is less virulent than Shiga-toxigenic E. coli having normal flagellin in a streptomycin-treated mouse model (Rogers et al., Reduced virulence of an fliC mutant of Shiga-toxigenic Escherichia coli O113:H21, Infect. Immun. 74: 1962-66 (2006)). However, neither Rogers et al. nor others have demonstrated that EHEC flagella are useful in a vaccine in bovine to reduce colonization of EHEC, or that flagella can be used in a vaccine to help protect bovine from EHEC colonization. Because there is a lack of an effective vaccine to prevent or reduce colonization of EHEC in bovine, there is a need for such a vaccine.